Spatial and temporal foraging patterns of Queensland fruit fly, Bactrocera tryoni (Froggatt) (Diptera: Tephritidae), for protein and implications for management

Fruit flies require protein for reproductive development and actively feed upon protein sources in the field. Liquid protein baits mixed with insecticide are used routinely to manage pest fruit flies, such as Bactrocera tryoni (Froggatt). However, there are still some gaps in the underpinning science required to improve the efficacy of bait spray technology. The spatial and temporal foraging behaviour of B. tryoni in response to protein was investigated in the field. A series of linked trials using either wild flies in the open field or laboratory-reared flies in field cages and a netted orchard were undertaken using nectarines and guavas. Key questions investigated were the fly's response to protein relative to: height of protein within the canopy, fruiting status of the tree, time of day, season and size of the experimental arena. Canopy height had a significant response on B. tryoni foraging, with more flies foraging on protein in the mid to upper canopy. Fruiting status also had a significant effect on foraging, with most flies responding to protein when applied to fruiting hosts. B. tryoni demonstrated a repeatable diurnal response pattern to protein, with the peak response being between 12:0016:00 h. Season showed significant but unpredictable effects on fruit fly response to protein in the subtropical environment where the work was undertaken. Relative humidity, but not temperature or rainfall, was positively correlated with protein response. The number of B. tryoni responding to protein decreased dramatically as the spatial scale increased from field cage through to the open field. Based on these results, it is recommend that, to be most effective, protein bait sprays should be applied to the mid to upper canopies of fruiting hosts. Overall, the results show that the protein used, an industry standard, has very low attractancy to B. tryoni and that further work is urgently needed to develop more volatile protein baits.

Strong resistance to phosphine in the rusty grain beetle, Cryptolestes ferrugineus (Stephens) (Coleoptera: Laemophloeidae): its characterisation, a rapid assay for diagnosis and its distribution in Australia

BACKGROUND: The recent development of very high resistance to phosphine in rusty grain beetle, Cryptolestes ferrugineus (Stephens), seriously threatens stored-grain biosecurity. The aim was to characterise this resistance, to develop a rapid bioassay for its diagnosis to support pest management and to document the distribution of resistance in Australia in 20072011. RESULTS: Bioassays of purified laboratory reference strains and field-collected samples revealed three phenotypes: susceptible, weakly resistant and strongly resistant. With resistance factors of > 1000 x , resistance to phosphine expressed by the strong resistance phenotype was higher than reported for any stored-product insect species. The new time-to-knockdown assay rapidly and accurately diagnosed each resistance phenotype within 6 h. Although less frequent in western Australia, weak resistance was detected throughout all grain production regions. Strong resistance occurred predominantly in central storages in eastern Australia. CONCLUSION: Resistance to phosphine in the rusty grain beetle is expressed through two identifiable phenotypes: weak and strong. Strong resistance requires urgent changes to current fumigation dosages. The development of a rapid assay for diagnosis of resistance enables the provision of same-day advice to expedite resistance management decisions. (c) 2012 Commonwealth of Australia. Published by John Wiley & Sons, Ltd.

Sublethal Exposure to Phosphine Decreases Offspring Production in Strongly Phosphine Resistant Female Red Flour Beetles, Tribolium castaneum (Herbst)

The red flour beetle is a cosmopolitan pest of stored grain and stored grain products. The pest has developed resistance to phosphine, the primary chemical used for its control. The reproductive output of survivors from a phosphine treatment is an important element of resistance development but experimental data are lacking. We exposed mated resistant female beetles to 0.135 mg/L of phosphine for 48 h at 25°C. Following one week of recovery we provided two non-exposed males to half of the phosphine exposed females and to half of the non-exposed control females. Females that had been exposed produced significantly fewer offspring than non-exposed females. Females that remained isolated produced significantly fewer offspring than both exposed females with access to males and non-exposed controls (P<0.05). Some females were permanently damaged from exposure to phosphine and did not reproduce even when given access to males. We also examined the additional effects of starvation prior to phosphine exposure on offspring production. Non-exposed starved females experienced a small reduction in mean offspring production in the week following starvation, followed by a recovery in the second week. Females that were starved and exposed to phosphine demonstrated a very significant reduction in offspring production in the first week following exposure which remained significantly lower than that of starved non-exposed females (P<0.05). These results demonstrate a clear sublethal effect of phosphine acting on the female reproductive system and in some individuals this can lead to permanent reproductive damage. Pest population rebound after a fumigation may be slower than expected which may reduce the rate of phosphine resistance development. The results presented strongly suggest that phosphine resistance models should include sublethal effects. © 2012 Ridley et al.

HPLC analysis of plant sterol oxidation products

Increased interest in the cholesterol-lowering effect of plant sterols has led to development of plant sterol-enriched foods. When products are enriched, the safety of the added components must be evaluated. In the case of plant sterols, oxidation is the reaction of main concern. In vitro studies have indicated that cholesterol oxides may have harmful effects. Due their structural similarity, plant sterol oxidation products may have similar health implications. This study concentrated on developing high-performance liquid chromatography (HPLC) methods that enable the investigation of formation of both primary and secondary oxidation products and thus can be used for oxidation mechanism studies of plant sterols. The applicability of the methods for following the oxidation reactions of plant sterols was evaluated by using oxidized stigmasterol and sterol mixture as model samples. An HPLC method with ultraviolet and fluorescence detection (HPLC-UV-FL) was developed. It allowed the specific detection of hydroperoxides with FL detection after post-column reagent addition. The formation of primary and secondary oxidation products and amount of unoxidized sterol could be followed by using UV detection. With the HPLC-UV-FL method, separation between oxides was essential and oxides of only one plant sterol could be quantified in one run. Quantification with UV can lead to inaccuracy of the results since the number of double bonds had effect on the UV absorbance. In the case of liquid chromatography-mass spectrometry (LC-MS), separation of oxides with different functionalities was important because some oxides of the same sterol have similar molecular weight and moreover epimers have similar fragmentation behaviour. On the other hand, coelution of different plant sterol oxides with the same functional group was acceptable since they differ in molecular weights. Results revealed that all studied plant sterols and cholesterol seem to have similar fragmentation behaviour, with only relative ion abundances being slightly different. The major advantage of MS detection coupled with LC separation is the capability to analyse totally or partly coeluting analytes if these have different molecular weights. The HPLC-UV-FL and LC-MS methods were demonstrated to be suitable for studying the photo-oxidation and thermo-oxidation reactions of plant sterols. The HPLC-UV-FL method was able to show different formation rates of hydroperoxides during photo-oxidation. The method also confirmed that plant sterols have similar photo-oxidation behaviour to cholesterol. When thermo-oxidation of plant sterols was investigated by HPLC-UV-FL and LC-MS, the results revealed that the formation and decomposition of individual hydroperoxides and secondary oxidation products could be studied. The methods used revealed that all of the plant sterols had similar thermo-oxidation behaviour when compared with each other, and the predominant reactions and oxidation rates were temperature dependent. Overall, these findings showed that with these LC methods the oxidation mechanisms of plant sterols can be examined in detail, including the formation and degradation of individual hydroperoxides and secondary oxidation products, with less sample pretreatment and without derivatization.

Plant secondary compounds and soil microbial processes in carbon and nitrogen cycling in relation to tree species

The aim of this study was to explore soil microbial activities related to C and N cycling and the occurrence and concentrations of two important groups of plant secondary compounds, terpenes and phenolic compounds, under silver birch (Betula pendula Roth), Norway spruce (Picea abies (L.) Karst) and Scots pine (Pinus sylvestris L.) as well as to study the effects of volatile monoterpenes and tannins on soil microbial activities. The study site, located in Kivalo, northern Finland, included ca. 70-year-old adjacent stands dominated by silver birch, Norway spruce and Scots pine. Originally the soil was very probably similar in all three stands. All forest floor layers (litter (L), fermentation layer (F) and humified layer (H)) under birch and spruce showed higher rates of CO2 production, greater net mineralisation of nitrogen and higher amounts of carbon and nitrogen in microbial biomass than did the forest floor layers under pine. Concentrations of mono-, sesqui-, di- and triterpenes were higher under both conifers than under birch, while the concentration of total water-soluble phenolic compounds as well as the concentration of condensed tannins tended to be higher or at least as high under spruce as under birch or pine. In general, differences between tree species in soil microbial activities and in concentrations of secondary compounds were smaller in the H layer than in the upper layers. The rate of CO2 production and the amount of carbon in the microbial biomass correlated highly positively with the concentration of total water-soluble phenolic compounds and positively with the concentration of condensed tannins. Exposure of soil to volatile monoterpenes and tannins extracted and fractionated from spruce and pine needles affected carbon and nitrogen transformations in soil, but the effects were dependent on the compound and its molecular structure. Monoterpenes decreased net mineralisation of nitrogen and probably had a toxic effect on part of the microbial population in soil, while another part of the microbes seemed to be able to use monoterpenes as a carbon source. With tannins, low-molecular-weight compounds (also compounds other than tannins) increased soil CO2 production and nitrogen immobilisation by soil microbes while the higher-molecular-weight condensed tannins had inhibitory effects. In conclusion, plant secondary compounds may have a great potential in regulation of C and N transformations in forest soils, but the real magnitude of their significance in soil processes is impossible to estimate.

Molecular Biomonitoring during Rhizoremediation of Oil-Contaminated Soil

Rhizoremediation is the use of microbial populations present in the rhizosphere of plants for environmental cleanup. The idea of this work was that bacteria living in the rhizosphere of a nitrogen-fixing leguminous plant, goat's rue (Galega orientalis), could take part in the degradation of harmful monoaromatic hydrocarbons, such as benzene, toluene and xylene (BTEX), from oil-contaminated soils. In addition to chemical (e.g. pollutant concentration) and physical (e.g. soil structure) information, the knowledge of biological aspects (e.g. bacteria and their catabolic genes) is essential when developing the rhizoremediation into controlled and effective bioremediation practice. Therefore, the need for reliable biomonitoring methods is obvious. The main aims of this thesis were to evaluate the symbiotic G. orientalis - Rhizobium galegae system for rhizoremediation of oil-contaminated soils, to develop molecular methods for biomonitoring, and to apply these methods for studying the microbiology of rhizoremediation. In vitro, Galega plants and rhizobia remained viable in m-toluate concentrations up to 3000 mg/l. Plant growth and nodulation were inhibited in 500 mg/l m-toluate, but were restored when plants were transferred to clean medium. In the greenhouse, Galega showed good growth, nodulation and nitrogen fixation, and developed a strong rhizosphere in soils contaminated with oil or spiked with 2000 mg/l m-toluate. The high aromatic tolerance of R. galegae and the viability of Galega plants in oil-polluted soils proved this legume system to be a promising method for the rhizoremediation of oil-contaminated soils. Molecular biomonitoring methods were designed and/or developed further for bacteria and their degradation genes. A combination of genomic fingerprinting ((GTG)5-PCR), taxonomic ribotyping of 16S rRNA genes and partial 16S rRNA gene sequencing were chosen for molecular grouping of culturable, heterogeneous rhizosphere bacteria. PCR primers specific for the xylE gene were designed for TOL plasmid detection. Amplified enzyme-coding DNA restriction analysis (AEDRA) with AluI was used to profile both TOL plasmids (xylE primers) and, in general, aromatics-degrading plasmids (C230 primers). The sensitivity of the direct monitoring of TOL plasmids in soil was enhanced by nested C23O-xylE-PCR. Rhizosphere bacteria were isolated from the greenhouse and field lysimeter experiments. High genetic diversity was observed among the 50 isolated, m-toluate tolerating rhizosphere bacteria in the form of five major lineages of the Bacteria domain. Gram-positive Rhodococcus, Bacillus and Arthrobacter and gram-negative Pseudomonas were the most abundant genera. The inoculum Pseudomonas putida PaW85/pWW0 was not found in the rhizosphere samples. Even if there were no ecological niches available for the bioaugmentation bacterium itself, its conjugative catabolic plasmid might have had some additional value for other bacterial species and thus, for rhizoremediation. Only 10 to 20% of the isolated, m-toluate tolerating bacterial strains were also able to degrade m-toluate. TOL plasmids were a major group of catabolic plasmids among these bacteria. The ability to degrade m-toluate by using enzymes encoded by a TOL plasmid was detected only in species of the genus Pseudomonas, and the best m-toluate degraders were these Pseudomonas species. Strain-specific differences in degradation abilities were found for P.oryzihabitans and P. migulae: some of these strains harbored a TOL plasmid - a new finding observed in this work, indicating putative horizontal plasmid transfer in the rhizosphere. One P. oryzihabitans strain harbored the pWW0 plasmid that had probably conjugated from the bioaugmentation Pseudomonas. Some P. migulae and P. oryzihabitans strains seemed to harbor both the pWW0- and the pDK1-type TOL plasmid. Alternatively, they might have harbored a TOL plasmid with both the pWW0- and the pDK1-type xylE gene. The breakdown of m-toluate by gram-negative bacteria was not restricted to the TOL pathway. Also some gram-positive Rhodococcus erythropolis and Arthrobacter aurescens strains were able to degrade m-toluate in the absence of a TOL plasmid. Three aspects of the rhizosphere effect of G. orientalis were manifested in oil-contaminated soil in the field: 1) G. orientalis and Pseudomonas bioaugmentation increased the amount of rhizosphere bacteria. G. orientalis especially together with Pseudomonas bioaugmentation increased the numbers of m-toluate utilizing and catechol positive bacteria indicating an increase in degradation potential. 2) Also the bacterial diversity, when measured as the amount of ribotypes, was increased in the Galega rhizosphere with or without Pseudomonas bioaugmentation. However, the diversity of m-toluate utilizing bacteria did not significantly increase. At the community level, by using the 16S rRNA gene PCR-DGGE method, the highest diversity of species was also observed in vegetated soils compared with non-vegetated soils. Diversified communities may best guarantee the overall success in rhizoremediation by offering various genetic machineries for catabolic processes. 3) At the end of the experiment, no TOL plasmid could be detected by direct DNA analysis in soil treated with both G. orientalis and Pseudomonas. The detection limit for TOL plasmids was encountered indicating decreased amount of degradation plasmids and thus, the success of rhizoremediation. The use of G. orientalis for rhizoremediation is unique. In this thesis new information was obtained about the rhizosphere effect of Galega orientalis in BTEX contaminated soils. The molecular biomonitoring methods can be applied for several purposes within environmental biotechnology, such as for evaluating the intrinsic biodegradation potential, monitoring the enhanced bioremediation, and estimating the success of bioremediation. Environmental protection by using nature's own resources and thus, acting according to the principle of sustainable development, would be both economically and environmentally beneficial for society. Keywords: molecular biomonitoring, genetic fingerprinting, soil bacteria, bacterial diversity, TOL plasmid, catabolic genes, horizontal gene transfer, rhizoremediation, rhizosphere effect, Galega orientalis, aerobic biodegradation, petroleum hydrocarbons, BTEX

Origin of leaf rust adult plant resistance gene Rph20 in barley

Rph20 is the only reported, simply inherited gene conferring moderate to high levels of adult plant resistance (APR) to leaf rust (Puccinia hordei Otth) in barley (Hordeum vulgare L.). Key parental genotypes were examined to determine the origin of Rph20 in two-rowed barley. The Dutch cultivar 'Vada' (released in the 1950s) and parents, 'Hordeum laevigatum' and 'Gull' ('Gold'), along with the related cultivar 'Emir' (a derivative of 'Delta'), were assessed for APR to P. hordei in a disease screening nursery. The marker bPb-0837-PCR, co-located with Rph20 on the short arm of chromosome 5H (5HS), was used to screen genotypes for the resistance allele, Rph20.ai. Results from phenotypic assessment and DNA analysis confirmed that Rph20 originated from the landrace 'H. laevigatum' (i.e., Hordeum vulgare subsp. vulgare). Tracing back this gene through the pedigrees of two-rowed barley cultivars, indicated that Rph20 has contributed APR to P. hordei for more than 60 years. Although there have been no reports of an Rph20-virulent pathotype, the search for alternative sources of APR should continue to avoid widespread reliance upon a single resistance factor.

Preservation of encapsulated shoot tips and nodes of the tropical hardwoods Corymbia torelliana × C. citriodora and Khaya senegalensis

Alginate encapsulation is a simple and cost-effective technique to preserve plant germplasm but there are only a few reports available on preservation of encapsulated explants of two highly valuable groups of tropical trees, the eucalypts (Myrtaceae) and mahoganies (Meliaceae). This study investigated alginate encapsulation for preservation of the eucalypt hybrid, Corymbia torelliana × C. citriodora, and the African mahogany, Khaya senegalensis. We assessed shoot regrowth of encapsulated shoot tips and nodes after storage for 0, 3, 6 and 12 months on media varying in sucrose and nutrient content, under storage conditions of 14°C and zero-irradiance. Encapsulated explants of both trees were preserved most effectively on high-nutrient (half-strength Murashige and Skoog) medium containing 1% sucrose, which provided very high frequencies of shoot regrowth (92–100% for Corymbia and 71–98% for Khaya) and excellent shoot development after 12 months’ storage. This technique provides an extremely efficient means for storage and exchange of eucalypts and mahoganies, ideally suited for incorporation into plant breeding and germplasm conservation programs.

Reduced fungicide use on a new Australian peanut cultivar, highly resistant to the late leaf spot and rust pathogens

Rust (caused by Puccinia arachidis) and late leaf spot (LLS, caused by Mycosphaerella berkeleyi) can cause significant yield losses in Australian peanut crops. Until recently, all commercial peanut varieties were highly susceptible to these pathogens, but the new Australian cultivar Sutherland has significantly higher levels of resistance than the older cultivars. Field trials were conducted at two sites in Queensland to (a) confirm the improved resistance of cv. Sutherland over another commercial cultivar, Menzies, (b) study the effects of timing of first spray, spray interval and cultivar on disease severity and yield, and (c) develop a suitable fungicide management program for cv. Sutherland. In the 2006 and 2007 trials, rust and LLS developed slower and had lower final disease ratings and AUDPC values on unsprayed plots of cv. Sutherland than on cv. Menzies. The timing of the first spray is critical in managing both rust and late leaf spot, with the results demonstrating that the first fungicide spray on cv. Sutherland should be applied as soon as rust and LLS are first seen on cv. Menzies. In most trials spray intervals of 14 days or 21 days were suitable to effectively control rust and LLS. In years with low disease pressure, few, if any, fungicide applications will be needed to manage the diseases, but in other years up to four sprays may be necessary. © Australasian Plant Pathology Society Inc. 2012.

Pheromone of the banana-spotting bug, Amblypelta lutescens lutescens Distant (Heteroptera: Coreidae): Identification, synthesis, and field bioassay

The banana-spotting bug, Amblypelta lutescens lutescens Distant (Heteroptera: Coreidae), is one of the principal pests of tree fruits and nuts across northern and eastern Australia. Apart from visual damage assessment, there are currently no reliable methods for monitoring bug activity to aid management decisions. An attractant pheromone for this species that could be used as a trap lure could potentially fill this void. Earlier, two male-specific compounds were identified in airborne extracts from A. lutescens lutescens, (E,E)-α-farnesene and (R,E)-nerolidol; an unknown compound with a molecular weight 220 was also detected. We now report the identification of this hitherto unknown compound as (R,E,E)-α-farnesene-10,11-oxide. Synthesis of this epoxide was conducted using a regioselective asymmetric dihydroxylation of a sulfolene. A blend mimicking the natural proportions of (E,E)-α-farnesene, (R,E)-nerolidol, and (R,E,E)-α-farnesene-10,11- oxide attracted male and female A. lutescens lutescens as well as nymphs in the field, verifying that the aggregation pheromone comprises or is contained within this group of compounds. Copyright © 2012 Ashot Khrimian et al.

The rph1 Gene Is a Common Contributor to the Evolution of Phosphine Resistance in Independent Field Isolates of Rhyzopertha Dominica

Phosphine is the only economically viable fumigant for routine control of insect pests of stored food products, but its continued use is now threatened by the world-wide emergence of high-level resistance in key pest species. Phosphine has a unique mode of action relative to well-characterised contact pesticides. Similarly, the selective pressures that lead to resistance against field sprays differ dramatically from those encountered during fumigation. The consequences of these differences have not been investigated adequately. We determine the genetic basis of phosphine resistance in Rhyzopertha dominica strains collected from New South Wales and South Australia and compare this with resistance in a previously characterised strain from Queensland. The resistance levels range from 225 and 100 times the baseline response of a sensitive reference strain. Moreover, molecular and phenotypic data indicate that high-level resistance was derived independently in each of the three widely separated geographical regions. Despite the independent origins, resistance was due to two interacting genes in each instance. Furthermore, complementation analysis reveals that all three strains contain an incompletely recessive resistance allele of the autosomal rph1 resistance gene. This is particularly noteworthy as a resistance allele at rph1 was previously proposed to be a necessary first step in the evolution of high-level resistance. Despite the capacity of phosphine to disrupt a wide range of enzymes and biological processes, it is remarkable that the initial step in the selection of resistance is so similar in isolated outbreaks.

Effect of Time of Planting and Plant Size on the Productivity of ‘Festival’ and ‘Florida Fortuna’ Strawberry Plants in a Subtropical Environment

The effect of time of planting and plant size on the performance of ‘Festival’ and ‘Florida Fortuna’ strawberry (Fragaria ×ananassa) plants was studied at Nambour in southeastern Queensland, Australia, over 2 years. The main objective of the work was to determine whether small plants yielded proportionally less than large plants as planting was delayed. First, bare-rooted transplants of ‘Festival’ were divided into small (crown diameters ranging from 6 to 10 mm) or large plants (10 to 17 mm) and planted in late March, mid-April, or late April. Second, transplants of ‘Florida Fortuna’ were divided into small (5 to 8 mm) or large plants (8 to 17 mm) and planted in early April, mid-April, or early May. The early planting for each cultivar corresponded with the time that the transplants are first available from commercial strawberry nurseries. Yields were generally greater in plants planted in late March/early April compared with plants planted later. Differences in yield between the small and large plants were consistent across the different times of planting, with the small plants always having lower yields. Small transplants are an issue for the productivity of strawberry fields in this environment whether they are planted early or late. Producers should consider paying a premium for large transplants delivered early in the season.

Mitochondrial DNA diversity of Cleruchoides noackae (Hymenoptera: Mymaridae): a potential biological control agent for Thaumastocoris peregrinus (Hemiptera: Thaumastocoridae)

Carpintero and Dellap, (Hemiptera: Thaumastocoridae) is a native Australian sap-feeding insect that has become invasive and seriously damaging to commercially grown in the Southern Hemisphere. Lin and Huber (Hymenoptera: Mymaridae) was recently discovered as an egg parasitoid of the Thaumastocoridae in Australia. Mitochondrial DNA (mtDNA; cytochrome oxidase subunit I, COI) sequence diversity amongst 104 individuals from these native populations revealed 24 sequence haplotypes. The COI haplotypes of individuals collected from the Sydney and Southeast Queensland clustered in distinct groups, indicating limited spread of the insect between the regions. Individuals collected from Perth in Western Australia were represented by four COI haplotypes. Although this population is geographically more isolated from other populations, two COI haplotypes were identical to haplotypes found in the Sydney region. The results suggest that has recently been introduced into Perth, possibly from the Sydney area. The high mtDNA diversity and limited spread that is suggested for is in contrast to the lack of geographic associated mtDNA diversity and extensive spread of . If implemented as a biological control agent, this factor will need to be considered in collecting and releasing .

Symptom development proceeds at different rates in susceptible and partially resistant cereal seedlings infected with Fusarium pseudograminearum

Variation in the reaction of cereal cultivars to crown rot caused by Fusarium spp., in particular Fusarium pseudograminearum, was identified over 50 yrs ago, however the parameters and pathways of infection by F. pseudograminearum remain poorly understood. Seedlings of wheat, barley and oat genotypes that differ in susceptibility to crown rot were inoculated with a mixture of F. pseudograminearum isolates. Seedlings were harvested from 7 to 42 days after inoculation and expanded plant parts were rated for severity of visible disease symptoms. Individual leaf sheaths were placed onto nutrient media and fungal colonies emerging from the leaf sheathes were counted to estimate the degree of fungal spread within the host tissue. Significant differences in both the timing and the severity of disease symptoms were observed in the leaf sheath tissues of different host genotypes. Across all genotypes and plant parts examined, the development of visible symptoms closely correlated with the spread of the fungus into that tissue. The degree of infection of the coleoptile and sub-crown internode varied between genotypes, but was unrelated to the putative resistance of the host. In contrast leaf sheath tissues of the susceptible barley cv. Tallon and bread wheat cv. Puseas scored higher disease ratings and consistently showed faster, earlier spread of the fungus into younger tissues than infections of the oat cv. Cleanleaf or the wheat lines 2-49 and CPI 133814. While initial infections usually spread upwards from near the base of the first leaf sheath, the pathogen did not appear to invade younger leaf sheaths only from the base, but rather spread laterally across from older leaf sheaths into younger, subtended leaf sheaths, particularly as disease progressed. Early in the infection of each leaf sheath, disease symptoms in the partially resistant genotypes were less severe than in susceptible genotypes, however as infected leaf sheaths aged, differences between genotypes lessened as disease symptoms approached maximum values. Hence, while visual scoring of disease symptoms on leaf sheaths is a reliable comparative measure of the degree of fungal infection, differences between genotypes in the development of disease symptoms are more reliably assessed using the most recently expanded leaf sheaths.